The overall goal of this proposal is to elucidate both the causes and effects of cannabinoid exposure on the biodistribution and metabolism of other drugs. A major aim will be to more fully characterize how cannabidiol (CBD)-pretreatment increases brain levels of other drugs of abuse and to determine the pharmacological consequences of those increases. CBD pretreatment not only markedly increases concentrations of tetrahydrocannabinol (THC) and its metabolites and metabolite half-times in the brain, but also increase the duration of THC-induced catalepsy. CBD pretreatment also increases brain levels of cocaine as well as acute cocaine-induced seizure activity and mortality. We will therefore determine the effect of CBD or THC pretreatment on brain levels of other drugs of abuse and its pharmacological consequences, as well as examine the effect of CBD or THC pretreatment on plasma protein binding, blood-brain barrier permeability, and circulating cytokines in order to characterize possible mechanisms contributing to the increased brain levels. Understanding the mechanisms involved in these alterations in brain pharmacokinetics is not only toxicologically important because marijuana is often coingested with many other drugs of abuse and clinically important drugs but could also prove advantageous, permitting increased entry into the CNS of specific pharmacological agents. Another specific objective includes an extension of our studies on the CBD-mediated modification of mouse P450 3A, by examining the effect of CBD on human P450 3A4. We therefore propose to isolate and characterize the CBD-modified peptide of human P450 3A4, as we have done for mouse P450 3A. Identification of the amino acid residue that is modified by CBD would increase our understanding of the active site of this clinically important P450, since direct x- ray crystallographic structural information of the enzyme active site is currently unavailable for this or any other mammalian P450. Site- directed mutagenesis studies will confirm the importance of this residue in catalytic function and in CBD-modification. Finally, we will determine the contribution of specific P450 subpopulations to the metabolism of cocaine in order to elucidate the mechanisms by which P450 bioactivates cocaine as well as to understand how CBD pretreatment protects against cocaine-induced liver damage. Our recent findings not only suggest that a presently unidentified P450(s) may contribute to cocaine bioactivation, but that it may also be susceptible to CBD-mediated inactivation.